Apoptosis is a form of directed cellular death that is essential for a variety of biological processes, including embryonic development, cancer surveillance, and host defense. Inappropriate apoptosis occurs in autoimmune disorders, malignancy, and acquired and heritable neurodegenerative disease. Two parallel pathways for apoptosis have been uncovered, one mediated by the release of cytochrome c from the mitochondrial intermembrane space, and a second that bypasses cytochrome c release by directly activating caspase 8. The release mechanism for cytochrome c remains controversial, with evidence suggesting that the mitochondrial outer membrane channels; VDACs, open to conduct cytochrome c. In mammals there exist three VDAC isoforms. The laboratory has generated cell lines and mice that are deficient for each VDAC or a subset of VDACs. It is proposed to use these cell lines and mice to define the role VDACs play in cytochrome c mediated apoptosis. Specifically, the kinetics and extent of apoptosis will be determined in the deficient cell lines and mice. Cytochrome c release will be directly quantified and binding of pro and anti-apoptotic protein factors to mutant mitochondria will be measured. Expression profiles following induction of apoptosis will be performed using microarrayed cDNAs. Finally, point mutations will be introduced into VDACs that will abrogate voltage dependent channel closure to determine if open channels interfere with apoptosis. These studies may validate VDACs as targets for inhibiting or enhancing apoptosis.